Visualization and phenotyping of proinflammatory antigen-specific T cells during collagen-induced arthritis in a mouse with a fixed collagen type II-specific transgenic T-cell receptor beta-chain

Introduction: The Vb12-transgenic mouse was previously generated to investigate the role of antigen-specific T cells in collagen-induced arthritis (CIA), an animal model for rheumatoid arthritis. This mouse expresses a transgenic collagen type II (CII)-specific T-cell receptor (TCR) b-chain and consequently displays an increased immunity to CII and increased susceptibility to CIA. However, while the transgenic Vb12 chain recombines with endogenous achains, the frequency and distribution of CII-specific T cells in the Vb12-transgenic mouse has not been determined. The aim of the present report was to establish a system enabling identification of CII-specific T cells in the Vb12-transgenic mouse in order to determine to what extent the transgenic expression of the CII-specific bchain would skew the response towards the immunodominant galactosylated T-cell epitope and to use this system to monitor these cells throughout development of CIA. Methods: We have generated and thoroughly characterized a clonotypic antibody, which recognizes a TCR specific for the galactosylated CII(260-270) peptide in the Vb12-transgenic mouse. Hereby, CII-specific T cells could be quantified and followed throughout development of CIA, and their phenotype was determined by combinatorial analysis with the early activation marker CD154 (CD40L) and production of cytokines. Results: The Vb12-transgenic mouse expresses several related but distinct T-cell clones specific for the galactosylated CII peptide. The clonotypic antibody could specifically recognize the majority (80%) of these. Clonotypic T cells occurred at low levels in the naïve mouse, but rapidly expanded to around 4% of the CD4 T cells, whereupon the frequency declined with developing disease. Analysis of the cytokine profile revealed an early Th1-biased response in the draining lymph nodes that would shift to also include Th17 around the onset of arthritis. Data showed that Th1 and Th17 constitute a minority among the CII-specific population, however, indicating that additional subpopulations of antigen-specific T cells regulate the development of CIA. Conclusions: The established system enables the detection and detailed phenotyping of T cells specific for the galactosylated CII peptide and constitutes a powerful tool for analysis of the importance of these cells and their effector functions throughout the different phases of arthritis. * Correspondence: [email protected] Section for Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Scheeles väg 2, 171 77 Stockholm, Sweden Full list of author information is available at the end of the article Merky et al. Arthritis Research & Therapy 2010, 12:R155 http://arthritis-research.com/content/12/4/R155 © 2010 Merky et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Introduction Collagen-induced arthritis (CIA) is the most commonly used animal model for rheumatoid arthritis. Development of CIA is dependent on both B cells and T cells. The major role of B cells is to produce collagen type II (CII)-specific antibodies, and passive transfer of such antibodies has the capacity to bind cartilage in vivo and induce an acute arthritis. A major role of T cells is to aid B cells in their production of anti-CII antibodies, but they are also believed to play an active part in the disease via activation of other cell types, such as synovial macrophages. The influence of T cells in established CIA, however, is less clear. Adoptive transfer of CII-specific T cells alone does not induce clinical disease but may lead to microscopic synovitis [1]. Adoptive transfer of CII-specific T cells has also been shown to prolong the otherwise acute arthritis induced by passive transfer of CII antibodies [2]. The use of T-cell receptor transgenic (TCR-tg) mice has proven a powerful tool for investigating the nature of self-reactive T cells in tolerance and autoimmunity [3]. To further facilitate the understanding for the role of T cells in CIA, three different CII-specific TCR-tg mouse strains have earlier been described and shown to display an accelerated onset of severe arthritis, compared with nontransgenic littermates. Transgenic T cells from all three strains are A-restricted and recognize the same region on CII that is located between amino acid positions 260 and 270. This region harbors a lysine residue at position 264, which is naturally subjected to post-translational modifications, through hydroxylation and subsequent glycosylation. Strikingly, each of the three previously described TCR-tg mouse strains in fact recognize different forms of the CII(260-270) epitope, where the Va11.1/Vb8.3-tg mouse [4], the Va11.1/ Vb8.2-tg mouse [5] and the Vb12-tg mouse [6] respond to the nonmodified [4], the hydroxylated [7] and the galactosylated [8] CII(260-270) peptide, respectively. Although each of the mentioned post-translationally modified peptides has its importance in A-restricted CIA, we have earlier shown that glycosylation of CII is of major importance for T-cell tolerance and pathology in CIA [9]. We therefore found it important to establish an animal model that would allow for identification and tracking of T cells specific for the galactosylated CII peptide. In contrast to the Va11.1/Vb8.3-tg and Va11.1/Vb8.2-tg mouse strains, however, which express both the a-chains and b-chains of the TCR as transgenes, the galactosylated CII-specific Vb12-tg mouse is only transgenic for the CII-specific b-chain, which may combine with any endogenous a-chain. Although the Vb12-tg mouse displays an increased T-cell and B-cell immunity to CII and is more susceptible to CIA, it cannot be assumed that all, or even the majority, of CD4 T cells are CII specific, as was the case for the Va11.1/Vb8.3-tg and Va11.1/Vb8.2-tg mouse strains. Indeed, the Vb12-tg mouse can also develop immunity against microorganisms as well as immunity to tuberculin-purified protein derivative, an immunogenic component of complete Freund’s adjuvant (CFA) [6,8]. In the present report we have therefore established and thoroughly characterized a clonotypic antibody, which recognizes a TCR specific for the galactosylated CII(260-270) peptide in the Vb12-tg mouse. Using this antibody we were able to describe the emerging T-cell response and its transition in Vb12-tg mice following challenge with CII. Materials and methods